Methods: A lentiviral vector expressing ILK-specific shRNA was constructed and introduced into cultured retinal Müller cells. Silencing of the ILK gene was identified by real time RT-PCR and Western blot. The Müller cell phenotype change was confirmed by immunodetection of alpha-smooth muscle actin (alpha-SMA) stress fiber formation. The generation of tractional force was assessed using a tissue culture assay with cells incubated in three-dimensional collagen gels; cell migration was determined by the Boyden chamber method, using 10% FBS as a chemotactic factor.

Mentions:
Phase-contrast images of primary cultured retinal Müller cells on the 3rd day after dissociation are shown in Figure 2A. Cells proliferated from isolated single cells or cell clusters and displayed two types of morphology; most of them were polygonal and flat, while a few were elongated. The cells were identified by staining for glutamine synthetase (GS), a specific marker of Müller cells. Müller cells are the only cells in the retina that express the enzyme glutamine synthetase (GS)28, 29, but expression of GS in cultured Müller cells is transient. Expression appears two days after dissociation and decreases when cells achieve confluence23. Figure 2B shows GS expression in cultured Müller Cells on the fourth day after dissociation. Positive GS staining was observed in the nuclei of small cells that appeared to be proliferating, while expression appeared to decrease in cells that were large and confluent.

Mentions:
Phase-contrast images of primary cultured retinal Müller cells on the 3rd day after dissociation are shown in Figure 2A. Cells proliferated from isolated single cells or cell clusters and displayed two types of morphology; most of them were polygonal and flat, while a few were elongated. The cells were identified by staining for glutamine synthetase (GS), a specific marker of Müller cells. Müller cells are the only cells in the retina that express the enzyme glutamine synthetase (GS)28, 29, but expression of GS in cultured Müller cells is transient. Expression appears two days after dissociation and decreases when cells achieve confluence23. Figure 2B shows GS expression in cultured Müller Cells on the fourth day after dissociation. Positive GS staining was observed in the nuclei of small cells that appeared to be proliferating, while expression appeared to decrease in cells that were large and confluent.

Methods: A lentiviral vector expressing ILK-specific shRNA was constructed and introduced into cultured retinal Müller cells. Silencing of the ILK gene was identified by real time RT-PCR and Western blot. The Müller cell phenotype change was confirmed by immunodetection of alpha-smooth muscle actin (alpha-SMA) stress fiber formation. The generation of tractional force was assessed using a tissue culture assay with cells incubated in three-dimensional collagen gels; cell migration was determined by the Boyden chamber method, using 10% FBS as a chemotactic factor.